Covered By Solapur University for B.Sc. III

1. Chemotherapy

2. Chemotherapy
Mr. Kamble Sainath Hanmant
Assistant Professor in Microbiology
D.B.F.Dayanand College of arts And Science,
Solapur

3. Antimicrobial Drugs
• The treatment of a disease with a chemical substance is known as
chemotherapy. The chemical substance is called a chemotherapeutic agent.
• An antimicrobial is an agent that kills microorganisms or inhibits their growth.
• Antimicrobial drugs are the chemotherapeutic drugs synthesized chemically.
• Antimicrobial medicines can be grouped according to the microorganisms they
act primarily against, For example, antibiotics are used against bacteria
and antifungal are used against fungi.
• They can also be classified according to their function.
• Agents that kill microbes are called microbicidal, while those that merely inhibit
their growth are called biostatic.
• The use of antimicrobial medicines to treat infection is known as antimicrobial
chemotherapy, while the use of antimicrobial medicines to prevent infection is
known as antimicrobial prophylaxis.

4. • In the late 1800s and early 1900s, when Paul Ehrlich was studying
microorganisms and how to stain them for microscopy, he imagined the idea
of a 'magic bullet' - a chemical that could specifically kill microbes but not
harm the host's own cells.
• Antimicrobial drugs are chemicals that are intended to have selective
toxicity against microbes, meaning that they kill microbial cells but not the
host's cells.
• Antimicrobial drugs include antibiotics, which were originally defined as
substances produced by microorganisms that inhibit other microorganisms.
• The name 'antibiotics' comes from the word 'antibiosis'.
• Unlike symbiosis, where two organisms live together in a way that is often
mutually beneficial, antibiosis is when one microorganism tries to kill
another one.

5. • Nowadays, the definition of antibiotics has changed a bit. When people say
'antibiotics,' they usually mean substances that inhibit bacteria.
• And instead of always being derived from microorganisms, many antibiotics
nowadays are made synthetically or semi-synthetically. This means that all or
part of the drug molecule is made by chemists in the lab.
• Antimicrobial drugs also include antifungal drugs, antiviral
drugs and antiparasitic drugs. These are chemicals that inhibit fungi, inhibit
viruses and inhibit parasites.
• Antimicrobials are drugs that destroy microbes, prevent their multiplication or
growth, or prevent their pathogenic action
• Differ in their physical, chemical, and pharmacological properties
• Differ in antibacterial spectrum of activity
• Differ in their mechanism of action

6. Antibiotics
• Antibiotics are chemical substances produced by various species of
microorganisms and other living systems that are capable of inhibiting the
growth of or killing bacteria and other micro organism.
• Antibiotics can be either
• Broad Spectrum – Kill a wide range of bacteria
• Narrow Spectrum – Kill a specific type or group of bacteria
• Antibiotics can be divided into two classes: Those antibiotics that kill
bacteria are called bactericidal, and those that merely suppress bacterial
growth are called bacteriostatic.
• Bacteriostatic antibiotics rely on the immune system to eradicate the non
multiplying bacteria from the patient.

7. • The susceptibility of a bacterial isolate to a given antibiotic is quantified by
the minimum inhibitory concentration (MIC) and the minimum bactericidal
concentration(MBC).
• As its name implies, the MIC measures the minimum concentration of
antibiotic that is still able to suppress growth of the bacterial isolate.
• MIC (minimal inhibitory concentration) the lowest concentration (highest
dilution) of the drug that has an inhibitory (bacteriostatic) effect.
• Likewise, the MBC is the minimum concentration of antibiotic that results
in killing of the bacterial isolate.
• MBC (minimal bactericidal concentration) the lowest concentration
(highest dilution) that has a killing (bactericidal) effect.

8. Properties of ideal Antimicrobial drugs
• 1) Selective Toxicity – Drug harms the microbe without causing significant
damage to the host.
• When searching for ways to treat disease, scientists look for differences
between the human (or animal) host and the pathogen.
• Ex. Penicillin interferes with cell wall synthesis. Animal cells have no cell
walls, so penicillin is not toxic to animals.
• 2) Spectrum of Activity – The range of different microbes against which an
antimicrobial agent acts.
• Example: Broad spectrum: Gram positive and Gram negative
bacteria vs. Narrow spectrum: Gram negative only.
• 3) Lack of “Side effects”. Antimicrobial drugs should not produce any
harmful side effects on the host cell.
• Toxicity – Some antimicrobials do exert toxic effects on the patients receiving
them.
• Ideal drugs do not exert toxic effects on the host cell.

9. Attributes of An Ideal Antimicrobial Drugs
• A. Solubility in body fluids
• B. Selective toxicity
• C. Toxicity not easily altered
(no food or drug interactions)
• D. Nonallergenic
• E. Stability (should be degraded and excreted by the body slowly)
• F. Resistance by microorganisms not easily acquired
• G. Long shelf life
• H. Reasonable cost

10. Mode of Action of following
Antimicrobial Drugs

11. Acting on Cell Wall: Penicillin
• Penicillin was the first naturally occurring antibiotic to be used for the
treatment of bacterial infections.
• Penicillin is one of group of compounds known as β- lactam antibiotics
which are unrivalled in the treatment of bacterial infections.
• The originally, this drug was obtained from the Penicillium fungi. The
original penicillins isolated directly from mold fermentations were mixtures
of compounds having different side chains.
• The addition of phenylacetic acid to the fermentation medium improved the
yield of penicillin and ensured that the product was substantially a single
compound known as penicillin G or benzylpenicillin.
• The first successful variant was obtained by replacing phenylacetic acid by
phenoxyacetic acid as the added precursor.

12. • Mode of Action
• Structurally, penicillins are β-lactam antibiotics.
• Bacterial cell walls are consisting of a protective peptidoglycan layer, which
is continuously undergoing remodeling.
• The remodelling process involves the breaking of the β-(1,4) linked N-
acetylmuramic acid and N-acetylglucosamine; as well as the breaking of the
cross-linking peptide chains.
• This cross-linking peptide chains is what provides the rigidity, to the otherwise
fluid cell wall.
• The breaking of this peptide cross-linking is performed by an enzyme called
transpeptidase.
• The transpeptidase also helps in reforming the peptide bonds once the
restructuring of the cell wall is done.
• The penicillins act by inhibiting this particular enzyme. By inhibiting this
enzyme the penicillin prevents the reformation of the peptide bonds and thus
makes the cell wall less strong.
• This loss of cell wall integrity causes the bacteria to leak out its cellular
contents and perish.

13. • This beta-lactam ring of the penicillin is generally not very stable and
therefore it participates in the inactivation of bacterial cell enzymes which
are essential for synthesis of peptidoglycan.
• Transpeptidase attacks the beta-lactam ring which opens up to give a more
stable compound.
• When this happens the drug remains bound to the transpeptidase via
covalent linkage and thereby inhibits the enzyme by acylation of the active
site.
• The resistance to penicillin arises due to mutations in the active site of the
transpeptidase enzyme.
• Thus there are many variants of the transpeptidase enzyme which need the
use of newer penicillin antibiotics.

14. • The penicillin- binding proteins or PBPs are regarded as the
specific targets for penicillin and the other β-lactam
antibiotics.
• As we shall see, the covalent reaction between β-lactam
antibiotics and the PBPs, which inactivates their
transpeptidase function but not the transglycosylase activity is
central to the antibacterial activity of these drugs.
• Penicillin binding proteins vary from species to species in
number, size, amount and affinity for β-lactams antibiotics.

15. Bacitracin
• Bacitracin is polypeptide antibiotic which is too toxic for systemic
administration but is sometimes used topically to kill Gram-positive bacteria
by interfering with cell wall biosynthesis.
• The antibiotic is ineffective against Gram- negative bacteria, probably
because its large molecular size hinders penetration through the outer
membrane to its target site.
• Bacitracin inhibits peptidoglycan biosynthesis by binding specifically to the
long chain C55 – isoprenol pyrophosphate in the presence of divalent metal
ions.
• In the formation of the linear peptidoglycan the membrane bound isoprenol
pyrophosphate is released.
• Normally this is converted by a pyrophosphatase to the corresponding
phosphate which thus becomes available for reaction with another molecule
of UDPMur-N-Ac-pentapeptide.

16. • Interaction between the lipid pyrophosphate and a metal ion- Bacitracin
coordination complex blocks this process and eventually halts the synthesis
of peptidoglycan.
• The identity of the divalent metal ion bound to the antibiotic in bacterial
cells in uncertain but could well be either Mg 2+ or Zn2+ .
• Bacitracin forms 1:1 complex with several divalent metal ions and
investigations employing nuclear magnetic resonance and optical rotatary
dispersion indicate the involvement of the imidazole ring of the histidine
residue of the antibiotic in metal ion binding .
• Additionally likely sites of metal ion interaction include the thiazoline
moiety and carboxyl groups of the D-aspartate and D- glutamate residue.

17. Vancomycin
• Vancomycin which is a member of a group of complex glycopeptide
antibiotics, was first isolated in the 1950s, but its real clinical importance
only emerged with the inexorable spread of methicillin- resistant
Staphylococci.
• The use of Vancomycin and structurally related glycopeptides has markedly
increased because of their value in treating serious infections caused by
MRSA and other Gram –positive bacteria.
• Because of their relatively large molecular size, the glycopeptides are
essentially inactive against the more impermeable Gram-negative bacteria.
• The antibacterial action of glycopeptide antibiotics depends on their ability
to bind specifically to the terminal D-alanyl –D-alanine group on the peptide
side chain of the membrane bound intermediates in peptidoglycan synthesis.

18. • It is important to note that this interaction occurs on the outer face of the
cytoplasmic membrane.
• The glycopeptide antibiotics probably do not enter the bacterial cytoplasm,
again because of their molecular size.
• The complex which is formed between vancomycin and D-alanyl D-alanine
blocks the transglycosylase involved in the incorporation of the
disaccharide- peptide into the growing peptidoglycan chain and DD-
transpeptidase and DD-carboxypeptidase for which the D-alanyl D-alanine
moiety is a substrate.
• Both peptidoglycan chain extension and cross linking are therefore inhibited
by glycopeptide antibiotics.

19. Acting on Protein Synthesis : Streptomycin
• This naturally occurring antibiotic is a member of the aminoglycoside group and
has the complex chemical structure.
• Streptomycin was discovered in the early 1940s and was the first drug really
effective against tuberculosis.
• It is broad spectrum antibiotic active against a range of Gram- positive and gram
negative bacteria.
• Its use limited by several problems, first the drug is effective only when given by
injection because its absorption from the gastrointestinal tract is very poor.
• Second along with others aminoglycosides, streptomycin may cause permanent
deafness.
• Streptomycin is bactericidal rather than bacteriostatic
• Cell death is preceded by marked effects on protein biosynthesis which are
specific for the 70s ribosomes of bacteria.

20. Structure of Streptomycin

21. • Mode of Action
• Streptomycin is a protein synthesis inhibitor.
• It binds to the small 16S rRNA of the 30S subunit of the bacterial ribosome,
interfering with the binding of formyl-methionyl-tRNA to the 30S subunit.
• This leads to codon misreading, eventual inhibition of protein synthesis and
ultimately death of microbial cells through mechanisms that are still not
understood.
• Speculation on this mechanism indicates that the binding of the molecule to the
30S subunit interferes with 50S subunit association with the mRNA strand.
• Streptomycin distorts the proofreading selection of the correct aminoacyl –tRN
A by the ribosome.
• Streptomycin strongly inhibits the initiation of peptide chains. The drug also
shows the elongation of partly completed chains, although even at high
concentration of streptomycin chain elongation is not completely suppressed.

22. • This results in an unstable ribosomal-mRNA complex, leading to
a frameshift mutation and defective protein synthesis; leading to cell
death.
• Humans have ribosomes which are structurally different from those
in bacteria, so the drug does not have this effect in human cells.
• At low concentrations, however, streptomycin only inhibits growth
of the bacteria by inducing prokaryotic ribosomes to misread mRNA.
• Ability of the aminoglycosides to induce ribosomal misreading of
mRNA is an important factor in their bactericidal action.
• Streptomycin is an antibiotic that inhibits both Gram-positive and
Gram-negative bacteria, and is therefore a useful broad-spectrum
antibiotic.

23. Chloramphenicol
• Chloramphenicol was originally derived from the bacterium Streptomyces venezuelae,
isolated by David Gottlieb, and introduced into clinical practice in 1949, under the
trade name Chloromycetin.
• Chloramphenicol is a naturally occurring antibiotic that is now entirely produced by
chemical synthesis.
• It was one of the first broad- spectrum antibiotics to be discovered and has excellent
bacteriostatic activity against both Gram-positive and Gram- negative cocci and
bacilli, as well as rickettsias, mycoplasmas and Chlamydia.
• Chloramphenicol has a broad spectrum of activity and has been effective in treating
ocular infections caused by a number of bacteria including Staphylococcus aureus,
Streptococcus pneumoniae, and Escherichia coli.
• Common side effects include bone marrow suppression, nausea, and diarrhea.The
bone marrow suppression may result in death. To reduce the risk of side effects
treatment duration should be as short as possible

24. Chemical structure of chloramphenicol

25. • Mode of action
• The bacteriostatic action of chloramphenicol is due to a specific
inhibition of protein biosynthesis on 70S ribosomes: it is completely
inactive against 80S ribosomes.
• Studies with radioactively labelled chloramphenicol show that it
binds exclusively to the 50S subunit to a maximum extent of one
molecule per subunit, the binding is completely reversible.
• Biochemical evidence strongly indicates that chloramphenicol
blocks peptide bond formation by inhibiting the peptidyl transferase
activity of the 50S subunit.
• It prevents protein chain elongation by inhibiting the peptidyl
transferase activity of the bacterial ribosome.

26. • It specifically binds to A2451 and A2452 residues in the 23S rRNA of the
50S ribosomal subunit, preventing peptide bond formation.
• While chloramphenicol and the macrolide class of antibiotics both interact
with ribosomes, chloramphenicol is not a macrolide.
• It directly interferes with substrate binding, whereas macrolides sterically
block the progression of the growing peptide.
• The most serious side effect of chloramphenicol treatment is aplastic
anaemia. This effect is rare and sometimes fatal.

27. Acting on nucleotide synthesis: Quinolones
• These compounds compose one of the most important groups of wholly
synthetic antibacterial drug in current medical use.
• Nalidixic acid and oxolinic acid are the so-called first genenreation
quinolones, whose spectrum of antibacterial action is confined to Gram-
negative bacteria.
• The introduction of fluorine atom at position C-6 in the second generation
compound ciprofloxacin resulted in a marked increase in potency and
extended the antibacterial spectrum to important Gram-positive pathogens.
• The quinolones are a family of synthetic broad-spectrum antibiotic drugs.
• The antibacterial activity of the quinolones is primarily due to inhibition of
DNA gyrase.
• When the isolated enzyme is incubated with DNA and a quinolone, the
supercoiling reaction is arrested at the point where the cut ends of the DNA
strands are covalently linked to the hydroxyl groups of the tyrosine-122
residues of GyrA.
• The re-ligation of the broken strands is blocked and supercoiling reaction can
be said to have been frozen midway.

28. • This results in the accumulation of double- stranded
nicks in the bacterial genome and may also prevent the
essential movements of DNA and RNA polymerase
along the DNA template.
• The bactericidal action of the quinolones probably
arises from a combination of these effects.
• The primary site of quinolone action is assigned to the
GyrA subunit because the most common mutations that
confer resistance to these drugs are found in a region of
GyrA referred to as the quinolone resistance
determining region or QRDR.
• Quinolones bind strongly to gyrase complexed with
DNA but only weakly to either the enzyme or DNA
alone

29. • Quinolones and fluoroquinolones inhibit bacterial replication by
blocking their DNA replication pathway.
• DNA is the core genetic material of the cells, and is responsible for
proper functioning of the cell.
• During protein synthesis and DNA replication, the double-stranded DNA
needs to unwind into a single stranded structure, which allows for
complementary base pairing to occur and synthesis of mRNA to procede.
• This unwinding of DNA in the bacteria is done by enzymes in the
bacteria called DNA gyrase or DNA topoisomerase.
• DNA gyrase is a topoisomerase II type enzyme that unwinds the DNA
by introducing negative supercoils and can also help relax positive
supercoils.
• Quinolones and fluoroquinolones inhibit this enzyme by binding to the
A-subunit of the enzyme due to which the bacteria is unable to replicate
or even synthesize proteins.

30. Rifampicin
• Rifampicin, also known as rifampin, is an antibiotic used to treat
several types of bacterial infections.
• Rifampicin is active against many Gram-positive bacteria, but less
effective against Gram- negatives because of limited access to the
target enzyme in these organisms.
• Rifampicin (Rif) is one of the most potent and broad spectrum
antibiotics against bacterial pathogens and is a key component of anti-
tuberculosis therapy, stemming from its inhibition of the bacterial RNA
polymerase (RNAP).
• Rifampicin may be given either by mouth or intravenously.
• Chemically the Rifampicin are closely related to the streptovaricins and
having similar mode of action.

31. • The transcription of RNA from DNA is one course, common
to both prokaryotic and eukaryotic organisms and involves
enzymes known as DNA-dependent RNA polymerase.
• The RNA polymerase isolated from E.coli is a large complex
consisting of four kinds of subunits: αββ’and σ.
• The complete or holoenzyme has the composition (α2ββ’σ)
together with two tightly bound Zinc atom.

32. • Rifampicin inhibits bacterial DNA-dependent RNA synthesis by
inhibiting bacterial DNA-dependent RNA polymerase.
• Crystal structure data and biochemical data suggest that rifampicin
binds to the pocket of the RNA polymerase β subunit within the
DNA/RNA channel, but away from the active site.
• The inhibitor prevents RNA synthesis by physically blocking
elongation, and thus preventing synthesis of host bacterial proteins.
• By this "steric-occlusion" mechanism, rifampicin blocks synthesis
of the second or third phosphodiester bond between the nucleotides
in the RNA backbone, preventing elongation of the 5' end of the
RNA transcript past more than 2 or 3 nucleotides.